The Section has continued to focus on structure-function studies of the noncollagenous proteins of bones and teeth, with particular emphases on matrix protein-protein and matrix protein-cell interactions. It is highly likely that both the assembly of the matrix and its subsequent mineralization are controlled by cells via the use of noncollagenous proteins including specific proteases. Other noncollagenous proteins are likely used in cell-cell and cell-matrix signal transduction. We have continued projects studying the roles that some of the ?bone and tooth? matrix proteins may play in normal ducts of the salivary gland and kidney as well as their roles in the development and metastasis of tumors. This year we have reached the intermediate goals in several areas discussed below. INTERACTION OF SIBLINGS AND MMPS IN NORMAL TISSUES (78% EFFORT). We have continued to present evidence to support our hypothesis that five small integrin-binding proteins [bone sialoprotein (BSP), osteopontin (OPN), dentin matrix protein 1 (DMP1), dentin sialophosphoprotein (DSPP) and matrix extracellular phosphoglycoprotein (MEPE)] whose genes are clustered within 375,000 base pairs on human chromosome 4 (mouse chromosome 5) are a gene family with related biological functions. We have named this family the SIBLINGs (Small Integrin-Binding LIgand, N-linked Glycoproteins.) One key element supporting the concept that these genes are likely the result of gene duplication and subsequent divergence is that we have recently shown that three members of the SIBLING family bind and activate three different members of the matrix metalloproteinase (MMP) family. All of the SIBLINGs are clearly expressed in bones and teeth and they were generally thought to be rarely expressed outside of the skeleton. Using monoclonal antibodies that we have recently generated against all five human SIBLING proteins, the Section is currently challenging this concept by documenting the presence of all five gene products in several normal, metabolically active epithelial tissues including kidney and salivary gland. For example, the SIBLINGs and their protein-cutting partners, the matrix metalloproteinases (MMP), are all expressed in specific ductal structures of human kidney. The different locations of each SIBLING and its MMP partner along the physiologically well-defined kidney tubule will help us hypothesize the possible roles that these proteins play in normal biology. We have theorized that these proteins play a critical role in the long-term maintenance of the ductal systems of metabolically active ductal epithelia. These proteins will also likely serve as useful identifiers of mature salivary gland ductal cells in our colleagues? current salivary gland stem cell cloning and other cell culture projects. Current projects include the activation of MMP-14 by cell surface BSP/MMP-2 complexes and the use of these complexes by normal cells for migration to new sites of matrix synthesis. MEPE is being investigated for a possible role in phosphate transport and DSPP in calcium transport. The expression of the SIBLING family members and their MMP partners in other metabolically active epithelial tissues remains an active research interest. SIBLING EXPRESSION IN CANCER (20% EFFORT) Our work with our collaborators in Belgium over the last 10 years has shown that various members of the SIBLING family of proteins are often turned on in the cells of many different types of tumors. This past work was based on immunohistology as well as quantitative serum assays. This year we used a cancer patient normal/tumor tissue array technology to survey the expression of the SIBLING gene products in a wide variety of cancers. The results not only verified that many different types of tumors express one or more members of the SIBLING family but also showed that there was often a coordinate expression of their partner MMP in the same patient?s tumor. For example, BSP mRNA expression was positively correlated with the message level of its protease partner, MMP-2, in breast and colon cancer. OPN correlated with MMP-3 in stomach and ovarian cancer while DMP1 correlated with MMP-9 in kidney and lung cancers. Furthermore, the expression levels of SIBLINGs were distinct within subtypes (e.g. breast ductal tumors compared to breast lobular tumors). In general, SIBLING expression increased with cancer stage for breast, colon, lung and rectal cancer. We conclude that SIBLINGs are potential markers of early cancer progression in a number of different cancer types, some of which currently lack vigorous clinical markers. Current projects include the use of SIBLINGs in precancerous and frank tumor diagnosis in head and neck cancers. We are also investigating the role of cell surface DMP1/MMP-9 complexes in the metastasis of some cancers. MAKING AND DISTRIBUTING ANTISERA AND OTHER PROBES (2% EFFORT) The Matrix Biochemistry Unit freely gives probes (antisera, cDNA, proteins etc.) to any laboratory in the world that makes a reasonable request. In FY05 we sent ~230 probes to 100 laboratories (27% were in the dental field) around the world. Others in the Branch have sent these same probes during this time as well.